幽门螺杆菌UreB与大肠杆菌LTB融合蛋白的表达及生物活性研究

Expression and Biological Activity of Fusion Protein of Hettcobacter pylori Urease B Subunit and E. coli Heat-labile Enterotoxin B Subunit

目的 通过基因工程方法构建并表达幽门螺杆菌尿素酶B亚单位(UreB)和大肠杆菌不耐热肠毒素B亚单位(LTB)以基因形式的融合蛋白,并对其生物活性进行初步研究,为幽门螺杆菌疫苗的研究奠定基础。方法 用PCR方法从本室构建的融合基因克隆载体扩增出融合基因2 040bp的片段,将融合基因插入原核表达载体pET-11-c中。结果 经全自动测序仪测序,SDS-PAGE和免疫印迹以及N端氨基酸测序分析,证实融合蛋白已在BL21中表达,初步纯化后通过动物实验和酶联免疫吸附试验证实其相应的免疫原性和免疫反应性,以及其中LTB成分和GM1结合的特性。结论 融合蛋白表达方式为幽门螺杆菌分子内佐剂疫苗的研究奠定了基础。

Objective To express and study the biological activity of fusion protein of Helicobacter pylori urease B subunit and E. coli heat-labile enterotoxin B subunit. Methods Amplify the fusion gene of Helicobacter pylori urease B subunit and E. coli heat-labile entertoxin B subunit from the constructed recombi-nant plasmid pFLU by PCR and insert into vector pET-11-c for expression in E. coli BL21. Results SDS-PAGE, Western blot and sequencing of amino acids at N-terminal proved that the fusion protein was successfully expressed. Animal test and ELBA showed good immunogenicity and immunoreactivity of the expressed product and the binding capacity of LTB component to GM1. Conclusion The expression of fusion protein laid a foundation of developing HP intramolecular adjuvant subunit vaccine.